乙炔抑制法在硝化与反硝化过程中的应用

硝化和反硝化作用在土壤氮素循环中扮演重要作用,由于硝化和反硝化作用一方面能够导致土壤中氮素的损失,另一方面能够产生温室气体-N2O,所以硝化和反硝化作用的研究备受关注.乙炔抑制法能同时测定硝化和反硝化作用,在硝化和反硝化作用中有着重要的应用.该文主要论述了乙炔抑制法的研究进展;以及对应用乙炔气体时存在的一些问题进行了综述.     

土地利用方式和培养温度对土壤氮转化及温室气体排放的影响

在好氧条件下研究土地利用方式(林地、草地)及培养温度(10、15 ℃)对加拿大和中国土壤的硝化作用、氮矿化作用以及N2O和CO2排放的影响.结果表明:草地土壤中的硝化作用和N2O排放量大于林地土壤,中国草地土硝化作用最强.10和15℃下中国草地土硝化速率分别为2.10和2.86 mg N·kg-1·d-1,15 d的N2O累积排放量分别为10.2和15.4μg N2O-N·kg-1.pH是影响土壤硝化作用强度和N2O排放的主要因素,与两者均呈显著正相关.林地土壤的矿化作用和CO2排放量高于草地,中国林地土壤的矿化作用最强,其平均矿化速率在10和15℃时分别为3.08和2.87mgN·kg-1 ·d-1.加拿大林地土壤CO2排放量最高,其15 d的累积排放量在10和15℃时分别为314和370 mg CO2-C·kg-1,土壤有机碳和水溶性有机碳含量分别与有机氮矿化作用和CO2排放量呈显著正相关.温度增加促进草地土壤硝化作用及林地和草地土壤中N2O的排放,也显著促进林地土壤中CO2的排放.     

与氮转化有关的土壤酶活性对抑制剂施用的响应

利用室内模拟培养试验,研究好气条件下施用尿素后土壤脲酶、脲酸还原酶、亚硝酸还原酶和羟胺还原酶活性对脲酶抑制剂氢醌（HQ）与硝化抑制剂包被碳化钙（ECC）和双氰胺（DCD）组合（HQ ECC、HQ DCD）的响应、结果表明,HQ DCD组合与其它抑制剂处理相比能更有效地降低土壤脲酶活性,增加硝酸还原酶、亚硝酸还原酶、羟胺还原酶活性,不同处理土壤脲酶、亚硝酸还原酶和羟胺还原酶活性与土壤NH4^ 、NO3^-、NH3挥发和N2O排放速率间存在不同形式的显著相关关系：土壤脲酶、亚硝酸还原酶和羟胺还原酶活性之间存在不同形式的显著正相关关系。     

真菌异化硝酸盐还原机理的研究进展

真菌异化硝酸盐还原途径的发现打破了反硝化仅存在于原核细胞这一传统观念。真菌异化硝酸盐还原途径是在环境中氧供给受限的情况下发生的, 包括反硝化和氨的发酵。硝酸盐能诱导产生反硝化作用的酶, 其中, 硝酸盐还原酶与亚硝酸还原酶位于线粒体中, 它们所催化的酶促反应能偶联呼吸链ATP合成酶合成ATP, 同时产生NO。与参与反硝化作用前两个酶不同, 真菌NO还原酶能以NADH为直接电子供体将NO还原为N2O, 在NAD+的再生和自由基NO的脱毒中起着重要作用。氨发酵则将硝酸盐还原成NH4+, 同时偶联乙酸的生成和底物水平磷酸化。此文从参与该过程的关键酶、关键酶的表达调节、真菌与细菌异化硝酸盐还原的比较等角度综述了真菌异化硝酸盐还原的最新研究进展。     

黄土性土壤剖面不同层次N2O浓度的原位监测

用土壤探头法对黄土性土壤玉米-小麦轮作体系下不同剖面层次N2O浓度变化进行了3a的田间原位监测.结果证实了黄土性土壤深层反硝化作用的存在,且N2O浓度有着明显的时间和空间变异.表现为N2O浓度受土壤气候条件(温度和降水)和生产管理措施的影响,丰水年明显高于亏水年;在降水或灌溉后出现瞬时N2O浓度峰.由于小麦和玉米生长特点和作物生长季气侯特点的差异,玉米生长季土壤剖面各层N2O浓度显著高于小麦生长期土壤剖面各层的浓度.统计分析结果表明:土壤剖面中不同土层N2O浓度的变化对照处理为60cm ≈ 90cm ≈150cm＞ 30cm＞ 10cm,而施肥处理为60cm > 90cm ≈150cm＞ 30cm＞ 10cm.深层土壤N2O的主要来源是土壤的反硝化作用,施肥显著地增加各土壤层次N2O的产生量.     

放牧家畜排泄物N转化研究进展

放牧家畜排泄物氮转化是草原生态系统氮循环的关键。自 2 0世纪 70年代以来 ,以提高氮利用效率和减少温室气体排放为目的的家畜排泄物氮转化的研究越来越受到人们的重视。放牧家畜排泄物氮的转化研究主要包括 3个方面 :氮的矿化、硝化与反硝化 ,氮的氨化。家畜粪氮矿化速度慢 ,持续时间长 ;尿氮矿化速度快 ,持续时间短。氮矿化与家畜排泄物 C∶ N比、木质素/氮素比、木质素含量和纤维素含量呈负相关关系 ,而与全氮含量和水溶性氮含量呈正相关 ;土壤动物和微生物可以显著促进氮的矿化过程 ;高温和相对干燥、砂质土壤较壤土和粘土有利于氮的矿化。 4～ 4 0℃氮硝化作用与温度呈正相关 ;硝化作用的底物和产物浓度、土壤溶液渗透压和氯化物浓度的增加对硝化作用有强烈的抑制效应 ;p H6 .0～ 8.0条件下硝化作用强度随着土壤p H值的升高而增加 ,而 p H值高于 8.0或低于 6 .0时硝化作用受到抑制 ;硝化作用与土壤氧气含量呈正相关关系 ,而与土壤含水量呈负相关 ;温暖湿润较干燥炎热的气候条件有利于硝化过程的进行。反硝化作用与土壤氧气浓度呈负相关关系 ,而与土壤含水量和可利用有机碳含量呈正相关 ;0～ 6 5℃反硝化作用强度随温度升高而增大 ,10～ 35℃条件下温度成为影响反硝化作用的关键因素 ;反硝化作用在     

CO_2浓度升高对水体硝化、反硝化作用的影响研究进展

大气CO_2浓度升高潜移默化地影响着水体生态系统的碳循环过程.然而,该过程如何影响与其耦合的氮循环过程仍不明确.水体硝化、反硝化过程作为水体氮循环的重要环节,必然会对大气CO_2浓度升高产生一系列的响应.本文总结了国内外关于大气CO_2浓度升高对水体理化性质、硝化作用、反硝化作用及N形态转化影响方面的研究工作,发现大气CO_2浓度升高会降低水体的p H,增加水中CO_2和HCO_3^-含量,但对富营养化与寡营养化水体中硝化、反硝化作用的影响具有明显差异.大气CO_2浓度升高抑制寡营养化水体的硝化作用和反硝化作用,降低N2_O的释放通量,抑制富营养化水体的硝化作用,但当水体pH在7～9时,可能促进反硝化作用,增加N2_O的释放通量,最终可能导致水体中NH_4^+的积累及NO_3^-浓度的降低,影响水体中微生物的多样性.在此基础上提出目前相关研究存在的瓶颈问题及值得深入探讨的科学问题,为进一步深入理解温室效应背景下全球CO_2浓度升高对水体生态系统N循环的影响提供参考.     

模拟咸水入侵对崇明岛河岸带根际土壤微生物及反硝化作用的影响

通过模拟咸水入侵,研究了其对崇明岛河岸带根际土壤微生物及其反硝化过程的影响.结果表明:模拟咸水入侵后4种不同植被型河岸带土壤根际微生物区系发生显著变化,除放线菌菌群数量稍有增加外,细菌、真菌以及硝化和反硝化细菌数量均出现不同程度下降,特别是反硝化功能细菌数量较对照平均下降51.8％,说明河岸带土壤不同微生物区系对咸水入侵的响应存在显著差异.模拟咸水入侵后,河岸带土壤与氮转换相关的酶活性普遍受到抑制,且抑制作用随酶类型不同而存在差异,亚硝酸还原酶对咸水入侵最敏感,其活性较对照平均下降了43.5％,脲酶活性次之,其降幅为37.4％,而脱氢酶受咸水影响较小,其活性平均下降29.5％.模拟咸水入侵明显削弱了河岸带土壤反硝化作用,其速率平均下降34.9％.不同植被型河岸带土壤微生物对咸水入侵的生态生理响应存在显著差异,与对照相比,茭白根际土壤微生物数量和酶活性受咸水入侵的平均抑制率最大,土壤反硝化速率最小,其次是菖蒲和芦苇.在模拟咸水入侵下,菖蒲与芦苇混合群落根际土壤微生物数量、酶活性和反硝化速率抑制率明显低于单一植物模式,表明混合植被群落根际土壤微生物过程及反硝化作用对咸水入侵具有较好的缓冲性能.     

牦牛排泄物输入对滇西北高寒泥炭沼泽湿地土壤氮转化的影响

牦牛放牧对滇西北高寒湿地土壤环境产生严重影响，改变土壤氮的迁移转化过程，影响湿地生态系统初级生产。然而，关于牦牛排泄物输入对滇西北高寒泥炭沼泽湿地土壤氮转化过程的影响尚不清楚。本研究以滇西北高原典型泥炭沼泽湿地为对象，采用原位土芯室内控制实验方法，研究牦牛排泄物输入对泥炭沼泽湿地土壤氮转化的影响。结果表明，粪便和尿液输入初期促进土壤铵态氮(NH₄⁺-N)积累，但整个培养期则表现为消耗NH₄⁺-N,积累硝态氮(NO₃^--N),表明该过程以硝化作用为主。粪便和尿液输入提高土壤脲酶活性(P<0.05),降低反硝化酶活性(P<0.05)。粪便输入提高过氧化氢酶活性(P<0.05)和N-乙酰氨基葡萄糖苷酶活性(P<0.05),尿液输入降低N-乙酰氨基葡萄糖苷酶活性(P<0.05)。粪便输入对土壤的矿化和硝化作用无显著性影响，尿液输入对土壤硝化作用影响显著。粪便输入抑制土壤反硝化作用，而尿液输入促进反硝化作用。牦牛排泄物输入通过影响泥炭沼...     

土壤释放的 N_2O 的原位测定

N_2O 是大气成分之一,它由微生物的硝化-反硝化作用、燃烧和大气闪电等过程产生,其中土壤微生物反硝化作用是最主要的来源。土壤微生物反硝化作用产生N_2O,不仅导致土壤中肥料氮素的损失,而且由于其“温室效应”和对臭氧层的破坏,受到国内外研究者     

Expression of denitrification enzymes in response to the dissolved oxygen level and respiratory substrate in continuous culture of Pseudomonas stutzeri

The onset and cessation of the synthesis of denitrification enzymes of Pseudomonas stutzeri were investigated by using continuous culture and defined dissolved oxygen levels covering the full range of transition from air saturation to complete anaerobiosis. Expression of nitrate reductase, nitrite reductase (cytochrome cd1), and N2O reductase was controlled by discrete oxygen levels and by the nature of the nitrogenous oxide available for respiration. N2O reductase was synthesized constitutively at a low level; for enhanced expression, oxygen concentrations were required to decrease below 5 mg of O2 per liter. The threshold values for synthesis of nitrate reductase and cytochrome cd1 in the presence of nitrate were ca. 5 and ca. 2.5 mg of O2 per liter, respectively. With nitrous oxide as the respiratory substrate, nitrite reductase was again the most sensitive to oxygen concentration; however, thresholds for all denitrification enzymes shifted to lower oxygen levels. Whereas the presence of nitrate resulted in maximum expression and nearly uniform induction of all reductases, nitrite and nitrous oxide stimulated preferably the respective enzyme catalyzing reduction. In the absence of a nitrogenous oxide, anaerobiosis did not induce enzyme synthesis to any significant degree. The accumulation of nitrite seen during both the aerobic-anaerobic and anaerobic-aerobic transition phases was caused by the differences in onset or cessation of synthesis of nitrate and nitrite reductases and an inhibitory effect of nitrate on nitrite reduction.     

Expression of denitrification enzymes in response to the dissolved oxygen level and respiratory substrate in continuous culture of Pseudomonas stutzeri.

The onset and cessation of the synthesis of denitrification enzymes of Pseudomonas stutzeri were investigated by using continuous culture and defined dissolved oxygen levels covering the full range of transition from air saturation to complete anaerobiosis. Expression of nitrate reductase, nitrite reductase (cytochrome cd1), and N2O reductase was controlled by discrete oxygen levels and by the nature of the nitrogenous oxide available for respiration. N2O reductase was synthesized constitutively at a low level; for enhanced expression, oxygen concentrations were required to decrease below 5 mg of O2 per liter. The threshold values for synthesis of nitrate reductase and cytochrome cd1 in the presence of nitrate were ca. 5 and ca. 2.5 mg of O2 per liter, respectively. With nitrous oxide as the respiratory substrate, nitrite reductase was again the most sensitive to oxygen concentration; however, thresholds for all denitrification enzymes shifted to lower oxygen levels. Whereas the presence of nitrate resulted in maximum expression and nearly uniform induction of all reductases, nitrite and nitrous oxide stimulated preferably the respective enzyme catalyzing reduction. In the absence of a nitrogenous oxide, anaerobiosis did not induce enzyme synthesis to any significant degree. The accumulation of nitrite seen during both the aerobic-anaerobic and anaerobic-aerobic transition phases was caused by the differences in onset or cessation of synthesis of nitrate and nitrite reductases and an inhibitory effect of nitrate on nitrite reduction.     

农田和森林土壤中氧化亚氮的产生与还原

采用土壤淤浆方法对丹麦农田和山毛榉森林土壤反硝化过程中Ｎ２Ｏ的产生与还原进行了研究。同时考察了硝酸根和铵离子对反硝化作用的影响。结果表明,森林土壤反硝化活性大于农田土壤,但农田土壤中Ｎ２Ｏ还原活性大于森林土壤,表现在农田和森林土壤中Ｎ２Ｏ／Ｎ２的产生比率分别为０．１１和３．６５。硝酸根和铵离子能促进两种土壤中的Ｎ２Ｏ产生,但可降低农田土壤中的Ｎ２Ｏ还原速率,与农田土壤相比,硝酸根可降低森林土壤Ｎ２     

Nitrous oxide production by Alcaligenes faecalis under transient and dynamic aerobic and anaerobic conditions.

Nitrous oxide can be a harmful by-product in nitrogen removal from wastewater. Since wastewater treatment systems operate under different aeration regimens, the influence of different oxygen concentrations and oxygen fluctuations on denitrification was studied. Continuous cultures of Alcaligenes faecalis TUD produced N2O under anaerobic as well as aerobic conditions. Below a dissolved oxygen concentration of 5% air saturation, the relatively highest N2O production was observed. Under these conditions, significant activities of nitrite reductase could be measured. After transition from aerobic to anaerobic conditions, there was insufficient nitrite reductase present to sustain growth and the culture began to wash out. After 20 h, nitrite reductase became detectable and the culture started to recover. Nitrous oxide reductase became measurable only after 27 h, suggesting sequential induction of the denitrification reductases, causing the transient accumulation of N2O. After transition from anaerobic conditions to aerobic conditions, nitrite reduction continued (at a lower rate) for several hours. N2O reduction appeared to stop immediately after the switch, indicating inhibition of nitrous oxide reductase, resulting in high N2O emissions (maximum, 1.4 mmol liter-1 h-1). The nitrite reductase was not inactivated by oxygen, but its synthesis was repressed. A half-life of 16 to 22 h for nitrite reductase under these conditions was calculated. In a dynamic aerobic-anaerobic culture of A. faecalis, a semisteady state in which most of the N2O production took place after the transition from anaerobic to aerobic conditions was obtained. The nitrite consumption rate in this culture was equal to that in an anaerobic culture (0.95 and 0.92 mmol liter-1 h-1, respectively), but the production of N2O was higher in the dynamic culture (28 and 26% of nitrite consumption, respectively).     

Denitrification by the fungus Cylindrocarpon tonkinense: anaerobic cell growth and two isozyme forms of cytochrome P-450nor.

We examined the denitrification system of the fungus Cylindrocapon tonkinense and found several properties distinct from those of the denitrification system of Fusarium oxysporum. C. tonkinense could form N2O from nitrite under restricted aeration but could not reduce nitrate by dissimilatory metabolism. Nitrite-dependent N2O formation and/or cell growth during the anaerobic culture was not affected by further addition of ammonium ions but was suppressed by respiration inhibitors such as rotenone or antimycin, suggesting that denitrification plays a physiological role in respiration. Dissimilatory nitrite reductase and nitric oxide reductase (Nor) activities could not be detected in cell extracts of the denitrifying cells. The Nor activity was purified and found to depend upon two isoenzymes of Cytochrome P-450nor (P-450nor), which were designated P-450nor1 and P-450nor2. These isozymes differed in the N-terminal amino acid sequence, isoelectric point, specificity to the reduced pyridine nucleotide (NADH or NADPH), and the reactivity to the antibody to P-450nor of F. oxysporum. the difference between the specificities to NADH and NADPH suggests that P-450nor1 and P-450nor2 play different roles in anaerobic energy acquisition.     

Physiological roles for two periplasmic nitrate reductases in Rhodobacter sphaeroides 2.4.3 (ATCC 17025)

The metabolically versatile purple bacterium Rhodobacter sphaeroides 2.4.3 is a denitrifier whose genome contains two periplasmic nitrate reductase-encoding gene clusters. This work demonstrates nonredundant physiological roles for these two enzymes. One cluster is expressed aerobically and repressed under low oxygen while the second is maximally expressed under low oxygen. Insertional inactivation of the aerobically expressed nitrate reductase eliminated aerobic nitrate reduction, but cells of this strain could still respire nitrate anaerobically. In contrast, when the anaerobic nitrate reductase was absent, aerobic nitrate reduction was detectable, but anaerobic nitrate reduction was impaired. The aerobic nitrate reductase was expressed but not utilized in liquid culture but was utilized during growth on solid medium. Growth on a variety of carbon sources, with the exception of malate, the most oxidized substrate used, resulted in nitrite production on solid medium. This is consistent with a role for the aerobic nitrate reductase in redox homeostasis. These results show that one of the nitrate reductases is specific for respiration and denitrification while the other likely plays a role in redox homeostasis during aerobic growth.     

Fungal denitrification and nitric oxide reductase cytochrome P450nor

We have shown that many fungi (eukaryotes) exhibit distinct denitrifying activities, although occurrence of denitrification was previously thought to be restricted to bacteria (prokaryotes), and have characterized the fungal denitrification system. It comprises NirK (copper-containing nitrite reductase) and P450nor (a cytochrome P450 nitric oxide (NO) reductase (Nor)) to reduce nitrite to nitrous oxide (N(2)O). The system is localized in mitochondria functioning during anaerobic respiration. Some fungal systems further contain and use dissimilatory and assimilatory nitrate reductases to denitrify nitrate. Phylogenetic analysis of nirK genes showed that the fungal-denitrifying system has the same ancestor as the bacterial counterpart and suggested a possibility of its proto-mitochondrial origin. By contrast, fungi that have acquired a P450 from bacteria by horizontal transfer of the gene, modulated its function to give a Nor activity replacing the original Nor with P450nor. P450nor receives electrons directly from nicotinamide adenine dinucleotide to reduce NO to N(2)O. The mechanism of this unprecedented electron transfer has been extensively studied and thoroughly elucidated. Fungal denitrification is often accompanied by a unique phenomenon, co-denitrification, in which a hybrid N(2) or N(2)O species is formed upon the combination of nitrogen atoms of nitrite with a nitrogen donor (amines and imines). Possible involvement of NirK and P450nor is suggested.     

土壤中反硝化酶活性变化与N2O排放的关系

研究施肥条件下,土壤反硝化酶活性硝酸还原酶(NR)活性、亚硝酸还原酶(NiR)活性及羟胺还原酶(HyR)活性在玉米生长季节中的变化及其与土壤含水量、硝态氮含量、N₂O排放之间的关系。结果表明,3种还原酶都有明显的季节变化规律并受土壤水分含量及施肥的影响。通过研究3种反硝化酶活性与土壤含水量及N₂O排放量之间的关系后指出,反硝化酶活性变化可作为一个区分旱田N₂O产生途径的指标.     

Nitrate reductase and respiratory adaptation in Bacillus stearothermophilus

Downey, R. J. (University of Notre Dame, Notre Dame, Ind.). Nitrate reductase and respiratory adaptation in Bacillus stearothermophilus. J. Bacteriol. 91:634-641. 1966.-Bacillus stearothermophilus 2184 required nitrate to grow in the absence of oxygen. Like many facultative microorganisms, the growth obtained anaerobically was considerably less than that obtained aerobically, even though the dissimilatory reduction of nitrate is, in effect, anaerobic respiration. The ability to reduce nitrate depended on the induction of nitrate reductase. Although oxygen at low levels did not retard induction of the enzyme, enzyme synthesis was considerably lessened by aeration. A semisynthetic medium containing nitrate supported aerobic growth of the thermophile but did not support anaerobic growth. The adaptation to nitrate resulted in a decrease in the level of cytochrome oxidase normally present in aerobically grown cells. Although the aerobic oxidation of succinate by the respiratory enzymes from aerobically grown cells was inhibited by 2-N-heptyl-4-hydroxyquinoline-N-oxide, the anaerobic oxidation of succinate by nitrate in a similar preparation from nitrate-adapted cells was not. The nitrate reductase in the bacillus was strongly inhibited by cyanide and azide but not by carbon monoxide. The nitrate reductase catalyzed the anaerobic oxidation of reduced nicotinamide adenine dinucleotide, and appeared to transfer electrons from cytochrome b(1) to nitrate. Cytochrome c(1) did not appear to be involved in the transfer.